Reddy A V, MacColl R, Maley F
Wadsworth Center for Laboratories and Research, New York State Department of Health, Albany 12201-0509.
Biochemistry. 1990 Mar 13;29(10):2482-7. doi: 10.1021/bi00462a007.
External invertase exists in an oligomeric equilibrium of dimer, tetramer, hexamer, and octamer, the concentrations of which vary with pH, time, and concentration of enzyme [Chu, F.K., Watorek, W., & Maley, F. (1983) Arch. Biochem. Biophys. 223, 543-555; Tammi, M., Ballou, L., Taylor, A., & Ballou, C.E. (1987) J. Biol. Chem. 262, 4395-4401]. To assess the influence of carbohydrate on this equilibrium, we investigated the self-association of external invertase (10 oligosaccharides per subunit), deglycosylated external invertase (2 oligosaccharides per subunit), and internal invertase (no carbohydrate) under various conditions. In addition, the effect of carbohydrate on the interaction of the subunits of these various invertases to form heterooligomers was studied. Chloride ion was found to promote subunit association in the various invertases irrespective of their glycosylation status. However, external invertase was less responsive to chloride ion relative to the internal and deglycosylated invertases. The higher oligomers of deglycosylated invertase were stable at 47 degrees C whereas those of external invertase dissociated rapidly into dimers, suggesting that the additional oligosaccharides in external invertase destabilize subunit interaction. Hybridization experiments with the various invertases showed that the dimers of internal invertase formed heterooligomers with either external or deglycosylated invertase. By contrast, the monomers of external and internal invertases formed their respective homodimers, but not heterodimers. These results suggest that the oligosaccharide content of invertase not only influences the extent of self-association but also affects heterooligomer formation.
外切转化酶以二聚体、四聚体、六聚体和八聚体的寡聚体平衡形式存在，其浓度随pH值、时间和酶浓度的变化而变化[朱，F.K.，瓦托雷克，W.，& 马利，F.（1983年）《生物化学与生物物理学报》223卷，543 - 555页；塔米，M.，巴卢，L.，泰勒，A.，& 巴卢，C.E.（1987年）《生物化学杂志》262卷，4395 - 4401页]。为了评估碳水化合物对这种平衡的影响，我们研究了外切转化酶（每个亚基10个寡糖）、去糖基化外切转化酶（每个亚基2个寡糖）和内切转化酶（无碳水化合物）在各种条件下的自缔合。此外，还研究了碳水化合物对这些不同转化酶亚基相互作用形成杂合寡聚体的影响。发现氯离子能促进各种转化酶中的亚基缔合，而不论其糖基化状态如何。然而，相对于内切转化酶和去糖基化转化酶，外切转化酶对氯离子的反应较弱。去糖基化转化酶的较高寡聚体在47℃时稳定，而外切转化酶的较高寡聚体则迅速解离成二聚体，这表明外切转化酶中额外的寡糖会破坏亚基间的相互作用。用各种转化酶进行的杂交实验表明，内切转化酶的二聚体与外切转化酶或去糖基化转化酶形成杂合寡聚体。相比之下，外切转化酶和内切转化酶的单体形成各自的同型二聚体，而不形成异型二聚体。这些结果表明，转化酶的寡糖含量不仅影响自缔合程度，还影响杂合寡聚体的形成。
